Project description:Cyanobacteria are oxygenic photoautotrophs responsible for a substantial proportion of nitrogen fixation and primary production in the hydrosphere. Non-nitrogen fixing cyanobacteria, such as Synechocystis sp. PCC 6803, depend of the availability of nitrogenized species to survive. Therefore, an intricate regulatory network around the transcriptional factor NtcA maintains the homeostasis of nitrogen in these organisms. The mechanisms controlling NtcA activity are well understood but a comprehensive study of its regulon is missing in Synechocystis. To define NtcA regulon during the early stage of nitrogen starvation, we have performed chromatin immunoprecipitation followed by sequencing (ChiP-seq), in parallel with genome level transcriptome analysis (RNA-seq). By combining both methods we assigned 51 activated and 28 repressed genes directly by NtcA. Most of direct targets included genes involved in nitrogen and carbon metabolism and photosynthesis. NtcA regulon also included 8 ncRNAs, of which ncr0710, Syr6 and NsiR7 were experimentally validated. Intriguingly, we identified several NtcA intragenic binding sites suggesting that NtcA can modulate transcriptional expression by binding along the whole transcript and not only in the promoter region as previously though. Finally, the transcriptional implication of PipX was analyzed in some NtcA-targets genes, revealing that PipX assists NtcA in the global nitrogen regulation in Synechocystis.
Project description:Cyanobacteria are oxygenic photoautotrophs responsible for a substantial proportion of nitrogen fixation and primary production in the hydrosphere. Non-nitrogen fixing cyanobacteria, such as Synechocystis sp. PCC 6803, depend of the availability of nitrogenized species to survive. Therefore, an intricate regulatory network around the transcriptional factor NtcA maintains the homeostasis of nitrogen in these organisms. The mechanisms controlling NtcA activity are well understood but a comprehensive study of its regulon is missing in Synechocystis. To define NtcA regulon during the early stage of nitrogen starvation, we have performed chromatin immunoprecipitation followed by sequencing (ChiP-seq), in parallel with genome level transcriptome analysis (RNA-seq). By combining both methods we assigned 51 activated and 28 repressed genes directly by NtcA. Most of direct targets included genes involved in nitrogen and carbon metabolism and photosynthesis. NtcA regulon also included 8 ncRNAs, of which ncr0710, Syr6 and NsiR7 were experimentally validated. Intriguingly, we identified several NtcA intragenic binding sites suggesting that NtcA can modulate transcriptional expression by binding along the whole transcript and not only in the promoter region as previously though. Finally, the transcriptional implication of PipX was analyzed in some NtcA-targets genes, revealing that PipX assists NtcA in the global nitrogen regulation in Synechocystis.
Project description:Elucidating the direct target regulon of NtcA during the early acclimation to nitrogen starvation in the cyanobacterium Synechocystis sp. PCC 6803 [ChIP-Seq]
Project description:Elucidating the direct target regulon of NtcA during the early acclimation to nitrogen starvation in the cyanobacterium Synechocystis sp. PCC 6803 [RNA-Seq]
Project description:The sRNA NsiR4 is involved in nitrogen assimilation control in cyanobacteria by targeting glutamine synthetase inactivating factor IF7
Project description:RNA-seq analysis of recombinant Synechocystis sp. 6803 provides important insights for understanding polyhydroxyalkanoate production in cyanobacteria under photoautotrophic conditions
Project description:Small proteins are an underinvestigated class of gene products in all domains of life. Here we describe the role of NsiR6/NblD, a cysteine-rich 66 amino acid small protein in the acclimation response of cyanobacteria to nitrogen starvation. Phycobilisomes, the macromolecular pigment-protein complexes for photosynthetic light harvesting, are rapidly degraded upon shift to low nitrogen. Deletion of nblD in Synechocystis sp. strain PCC 6803 prevents this degradation, indicated by the non-bleaching (nbl) phenotype. Complementation by a plasmid-localized gene copy fully restored the phenotype of the wild type, while overexpression of NblD under nitrogen-replete conditions did not lead to any phenotypical effect, different from the unrelated proteolysis adaptors NblA1 and NblA2, which can trigger phycobilisome degradation ectopically. However, transcriptome analysis revealed that nitrogen starvation induced nblA1/2 transcription in the ΔnblD strain, which excluded the possibility that the nbl phenotype was due to a possible NblD function as transcriptional co-regulator. In contrast, fractionation experiments indicated the presence of NblD in the phycobilisome fraction and pull-down experiments with NblD containing a triple FLAG tag identified the α and β phycocyanin subunits as the only two co-purifying proteins. Homologs of NblD exist in all cyanobacteria that use phycobilisomes but not in the genera Prochlorococcus and Acaryochloris which use alternative light-harvesting mechanisms. These data suggest that NblD plays a crucial role in the coordinated dismantling of phycobilisomes when nitrogen becomes limiting. We performed a microarray, to examine the global expression pattern of wild type and ∆nblD isolated RNA after 0h and 3h past induction of nitrogen depletion (-N) to detect potential differences in the nitrogen acclimation process.